Blastomycosis, a human systemic fungal disease caused by the organism Blastomyces dermatitidis, exists primarily in the southeastern and south central United States. Infection results from the inhalation of the fungal spore into the lung and a disease similar to tuberculosis is produced. The immunological diagnosis of blastomycosis is extremely difficult at the present time due to the unavailability of reliable antigens and the lack of specificity and sensitivity of the serological assays for serum antibodies. During the past three years our laboratory has developed sensitive procedures for the detection of immune responses associated with the fungal diseases histoplasmosis and coccidioidomycosis, which are similar to blastomycosis. It is proposed that this technique, the enzyme-linked immunosorbent assay (ELISA), will be adapted and optimized for the immunodiagnosis of blastomycosis. Yeast-phase antigens will be prepared from several strains of B. dermatitidis and compared with commercially available Blastomyces complement-fixation/immunodiffusion antigens using electrophoretic separation techniques and immunoblotting methods in an effort to determine antigenic composition. Studies will then be performed on purification, optimization and utilization of these antigens in ELISA and DOT-ELISA procedure to detect antibodies (IgG, IgM, IgA) in sera from patients with blastomycosis and other systemic fungal diseases. In addition comparative evaluations will be performed using different enzyme systems, different substrates and the avidin-biotin system in an effort to develop a highly specific and sensitive assay for the definitive diagnosis of blastomycosis. This proposed research work has the potential for future expansion to include (1) use of the antigens as skin-testing reagents to detect delayed dermal hypersensitivity (2) use of the reagents to produce polyclonal/monoclonal antibodies that could be used in ELISA procedures to detect antigens and (3) use of the antigens in vaccine preparations for the prevention of blastomycosis.